The objective of this research is to use insights from human genetics to define the molecular basis of disease in amyotropic lateral sclerosis (ALS), an incurable and fatal neurodegenerative disorder. The identification of causative mutations in TDP-43, along with the discovery of TDP-43 aggregate pathology, has shifted our understanding of the pathogenesis of ALS, and is a significant new direction in the field of neurodegenerative disease research. Ultimately, we would like to understand whether TDP-43 mutations lead to motor neuron degeneration through a toxic gain of function or a loss of normal functioning, arising from disruption of its normal roles as an RNA binding protein. Cellular and animal models are essential to understand the normal role of TDP-43, and the link between the mutations in the protein and neurodegeneration. In this grant, we propose to characterize a new mouse model of TDP-43 which will be invaluable in studying mechanisms of disease, and for investigating new therapeutic strategies for ALS. The transgenic mice we have generated express disease mutant TDP-43 throughout the nervous system, and develop a progressive and fatal neurodegenerative disease with features of ALS/FTLD-U (frontotemporal lobar degeneration with ubiquitinated inclusions). This proposal outlines experiments to determine the extent to which this mouse model recapitulates human ALS, and will define outcome measures that can be used in future translational studies, by performing detailed behavioral, biochemical, and pathologic analysis. To determine if altered mRNA splicing is a feature of the disease and to identify TDP-43 splicing targets, an exon-array will be performed on cortex of transgenic mice. A novel splicing dual-reporter assay will be used to directly examine the effect of wild-type and disease mutant forms of TDP-43 on splicing of exons known to be regulated by TDP-43, and to validate splicing targets identified by the exon-array. Together, these cellular and animal models will contribute to the understanding of the molecular mechanism of neurodegeneration from TDP-43 mutations. PUBLIC HEALTH RELEVANCE: Amyotrophic lateral sclerosis (ALS) is a devastating and fatal neurodegenerative disorder, for which no effective therapies exist. Mouse models of human disease are useful for understanding what causes neurodegenerative diseases, as well as for the development of new therapeutic agents. In this study, we will analyze a new mouse model that we have created which reflects underlying pathways that lead to ALS from TDP-43 mutations. Additionally, we will examine whether abnormalities in RNA processing plays a fundamental role in TDP-43 related ALS, in an effort to define new targets for therapeutic intervention in ALS.